Evidence for the signaling role of methyl jasmonate, methyl salicylate and benzothiazole between poplar (Populus simonii × P. pyramidalis ‘Opera 8277’) cuttings

Interplant communication has been widely demonstrated in plants, especially in herbaceous plants. In this study, mechanical damage was shown to affect the levels of pyrochatechol, chlorogenic acid, gallic acid and p-hydroxyl benzoic acid in poplar (Populus simonii × P. pyramidalis ‘Opera 8277’) cuttings, indicating the activation of defense response. In neighboring intact cuttings, the levels of those phenolics also varied when compared to the control, suggesting the interplant communication between poplar cuttings. Three volatiles, methyl jasmonate, methyl salicylate and benzothiazole, were detected in volatiles emitted from mechanically damaged poplar cuttings. All of them can induce changes in the levels of four phenolics. Therefore, they could act as airborne signals between P. simonii × P. pyramidalis ‘Opera 8277’ cuttings. The different change patterns of phenolic contents induced by different volatiles imply that the defense response activated in neighboring plants may be regulated by multiple signaling pathways. The results also suggest that the entire defense response of plants involves a variety of airborne signals in wound-induced volatiles.     

Correction of chromosomal point mutations in human cells with bifunctional oligonucleotides.

A sequence-specific genomic delivery system for the correction of chromosomal mutations was designed by incorporating two different binding domains into a single-stranded oligonucleotide. A repair domain (RD) contained the native sequence of the target region. A third strand-forming domain (TFD) was designed to form a triplex by Hoogsteen interactions. The design was based upon the premise that the RD will rapidly form a heteroduplex that is anchored synergistically by the TFD. Deoxyoligonucleotides were designed to form triplexes in the human adenosine deaminase (ADA) and p53 genes adjacent to known point mutations. Transfection of ADA-deficient human lymphocytes corrected the mutant sequence in 1-2% of cells. Neither the RD or TFD individually corrected the mutation. Transfection of p53 mutant human glioblastoma cells corrected the mutation and induced apoptosis in 7.5% of cells.     

Biosynthesis of rat insulin-like growth factor II. I. Immunochemical demonstration of a approximately 20-kilodalton biosynthetic precursor of rat insulin-like growth factor II in metabolically labeled BRL-3A rat liver cells

BRL-3A rat liver cells synthesize mature 7484-dalton rat insulin-like growth factor II (rIGF-II) as a approximately 22-kDa precursor, presumably prepro-rIGF-II. In the present study, we have biosynthetically labeled intact BRL-3A cells with [35S]cysteine and immunoprecipitated cell lysates and media with antisera to rIGF-II. A approximately 20-kDa protein was identified in immunoprecipitates of cell lysates having properties consistent with pro-rIGF-II. The approximately 20-kDa protein is precipitated by immune sera but not by nonimmune serum. Its immunoprecipitation is specifically inhibited by unlabeled rIGF-II but not by insulin. It is not precipitated from labeled lysates of a subclone of BRL-3A cells (BRL-3A2) that does not synthesize rIGF-II. The approximately 20-kDa protein is rapidly labeled intracellularly (10 min) but is not detected in BRL-3A media. In pulse-chase experiments, radioactivity in the approximately 20-kDa protein disappears during the chase and appears, at later times, in specifically immunoprecipitated approximately 19-, approximately 10-, approximately 8-, and approximately 7-kDa proteins in media and, to a limited extent, intracellularly. A protein with electrophoretic mobility identical to that of the approximately 20-kDa protein observed in cell lysates is immunoprecipitated from 35S-proteins whose synthesis is directed by BRL-3A RNA in a reticulocyte lysate cell-free translation system supplemented with microsomal membranes, and presumably arises by cotranslational removal of the signal peptide from approximately 22-kDa prepro-rIGF-II. Processing of the approximately 20-kDa protein in intact BRL-3A cells to intermediate and mature rIGF-II species appears to occur at the time of secretion and/or shortly thereafter, with the different forms appearing at approximately the same time.     

Methods for rapid screening and isolation of bacteria producing acidic lipase: feasibility studies and novel activity assay protocols

Acidic lipase finds its commercial values in medical applications and bioremediation of food wastes. In this work, approaches for rapid screening of lipase-producing bacteria were developed and the feasibility assessment of the screening methods was performed. From food waste samples, the proposed screening procedures allowed isolation of sixteen pure bacterial strains expressing higher lipase activity at acidic pH (pH 6.0) than at alkaline pH (pH 9.0). To enhance the accuracy of lipase activity determination under acidic conditions, a novel assay procedure was also developed by deactivating lipase activity by microwave treatment prior to back titration. This additional step could minimize interferences arising from residual lipase activity during conventional direct back-titration methods in measuring lipase activity at acidic pH. Using the four strategies proposed in this work, the best acidic-lipase-producing isolate was obtained by strategy C (SSC) and was identified as Aeromonas sp. C14, displaying an optimal lipase activity of 0.7 U/ml at an acidic pH of 6.0.     

An interspecific somatic hybrid between Actinidia chinensis and Actinidia kolomikta and its chilling tolerance

Protoplasts isolated from cotyledon-derived callus of Actinidia chinensisPlanch. var. chinensis (2N=2x=58) were fused with mesophyll protoplasts of Actiniadia kolomikta(Maxim. et Rupr.) Maxim (2N=2x=58) using a PEG method. Plantlets were regenerated from the fusion product clone 11. RAPD analyses, chromosome numbers of root tip cells and fluorescence peak position of leaf nuclei confirmed that clone 11 was an interspecific somatic hybrid (2N=4x=116) between A. chinensis and A. kolomikta. The chilling tolerance of the somatic hybrid was tested with in vitro leaves at low temperatures. Based on data of leaf thickness, electroconductivity, proline levels, malondialdehyde content and activity of superoxide dismutase, dendrogram cluster analysis suggested that the interspecific somatic hybrid was similar to A. kolomikta, and might have a higher capacity of cold resistance than A. chinensis.     

Inducible nuclear expression of NF-kappa B in primary B cells stimulated through the surface Ig receptor

Constitutive expression of NF-kappa B has been associated with developmental maturity in B cells on the basis of studies using continuously growing cell lines and plasmacytomas; however, little is known about the behavior of NF-kappa B in primary, mature B cells. In the present work, the regulation of NF-kappa B expression was studied by analyzing subcellular fractions of adult murine splenic B cells with the electrophoretic mobility shift assay using a kappa B-containing oligonucleotide. Although nuclear extracts from resting B cells contained kappa B-binding activity, additional kappa B-binding activity was present in cytosolic fractions in a form that became apparent after treatment with detergent. Competition analysis indicated that the DNA binding activity detected by electrophoretic gel mobility shift assay was specific for the kappa B motif, and UV photo-cross-linking showed the molecular size of kappa B-binding protein to be similar to that of the DNA binding subunit of NF-kappa B. Nuclear expression of kappa B-binding activity was markedly induced by treatment of B cells with phorbol ester or with LPS. Most notably, kappa B-binding activity was induced after surface IgR cross-linking, and the mechanism of this induction involved PKC. Further, anti-Ig-induced activity was superinduced in the presence of cycloheximide. These results indicate that nuclear NF-kappa B is rapidly induced as a result of B cell stimulation, and further suggest that NF-kappa B may play a specific role in mature B cells after ligand binding to surface Ig distinct from its postulated developmental role as a stage-specific factor involved in kappa-enhancer function.